learn more Moreover, in small lesions or advanced diseases, the possibility for retrieval of several biopsies can be limited. One study described the influence of the size of the biopsy needle in rat liver biopsies on the RNA quality in a subsequent micro-array expression study [12]. The aim of our study

was to assess different sampling techniques (with the optimal needle size as described above), fixation methods, and storage procedures for canine liver tissue. Our objective was to optimize the use of a single liver biopsy, in order to minimize the number of necessary biopsies per patient, by evaluation of different BIIB057 in vivo methods for RNA isolation and fixation available in our laboratory. Three biopsy techniques (wedge biopsy, Menghini, and True-cut), four storage methods for retrieval of RNA (snap freezing, RNAlater, Boonfix, RLT-buffer), two RNA isolation procedures (Trizol and RNAeasy), and three different fixation protocols for histological studies (10% formalin, RNAlater, Boonfix) were compared. Histological evaluation was based on hematoxylin-eosin (HE) and reticulin (fibrogenesis) staining, and rubeanic acid and rhodanine stains for copper.

KU55933 datasheet Immunohistochemical evaluation was performed for three different proteins at different (sub)cellular locations keratin-7 (K-7), multidrug resistance binding protein-2 (MRP-2) and Hepar-1. Results RNA isolation: RNAeasy mini kit versus Trizol The A260/A280 ratios of all samples in this study were between 1.98 and 2.13. The RNAeasy mini kit isolation was compared to the Trizol mediated isolation protocol in RNAlater fixed Menghini biopsies. RNA-quality of RNA isolated with the RNAeasy mini kit was consistently superior (1 to 1.5 RIN-values higher) to RNA isolated with the Trizol method (Table 1). Results from assessment of RNA quality prompted us to restrict further comparisons of different RNA fixation protocols to RNA isolated with the RNAeasy mini kit. Table 1 RIN-values after RNA isolation with RNAeasy mini kit or Trizol method (data of three independent representative isolations). RNAeasy

Trizol 8.1 7.3 8.8 7.4 8.2 6.7 Biopsy was taken with True-cut technique, RNA was stored in RNAlater. Independent samples were split and divided over the two isolation Vildagliptin procedures. Tissue fixation for RNA isolation RNA quality was compared between four methods of biopsy fixation: snap-freezing, Boonfix, B-RLT medium, and RNAlater. Table 2 depicts a comparison for RNA quality after RNA isolation with the RNAeasy mini kit. Three independent results per fixation protocol were measured. Snap-freezing, B-RLT, and RNAlater revealed RIN-values consistently within the range required for micro-array (range 7.9 to 9.3). A slight tendency for higher RIN-values for blind biopsies compared to True-cut biopsies. Since the RNA isolated from liver tissue fixed in Boonfix had RIN-values often below 8 (range 7.1–8.